The long-term goal of the Marsh-Armstrong laboratory is to discover new biological processes that may some day become treatment targets in glaucoma. Our laboratory discovered that astrocytes in the optic nerve head, the site where axon loss likely happens in glaucoma, are intrinsically phagocytic, and recently, that axonal mitochondria are one of the major targets of their phagocytosis. Preliminary studies of one the phagocytic genes expressed by optic nerve astrocytes, which is called Mfge8, have lead to the discovery that during development, injury and disease, but to some extent also normal adult physiology, there is much debris within the optic nerve head that is partially processed by astrocytes but then cleared through glymphatic channels around blood vessels that are contiguous with the subarachnoid space. In this application, we set out to examine these ?Glymphatics and Debris Clearance in Glaucoma?, to determine how the glymphatic system works at the optic nerve head and how it is affected in animal models of glaucoma. Many of the experiments will be carried out in mice, to: 1) determine the relationship between debris, glymphatic channels, and myeloid cells, which we propose enter the optic nerve center under conditions where debris is not properly removed, 2) determine how the glymphatics, debris clearance, and myeloid cells are affected in mouse models of glaucoma, and 3) determine what happens when debris builds up due to inhibition of the key phagocytic pathways. Other experiments will be carried out in a non-human primate model of glaucoma to: 1) determine whether glymphatics and debris clearance from the optic nerve are similar in mice and primates, and 2) determine whether they are similarly affected by glaucomatous insults. The primate studies will use some of the same tools used in the mouse studies, to be able to integrate the results coming from both species, but also very high resolution reconstructions of areas within the optic nerve that have low amounts of axon damage, that is, areas representing early stages in glaucomatous damage. Altogether, the proposed studies will rigorously test the idea that efficient debris clearance by astrocytes is critical for maintaining a healthy optic nerve, and that in the absence of efficient debris clearance there is myeloid infiltration and axonal damage. Should the experiments demonstrate that the proposed hypothesis is largely correct, it would represent a significant departure from current views of how glaucomatous axonal loss occurs, and this would have important implications as to how glaucoma should be diagnosed and treated.